The development of 11 C-carbonylation chemistry: A systematic view

The prospects for using carbon-11 labelled compounds in molecular imaging has improved with the development of diverse synthesis methods, including 11 C-carbonylations and re ﬁ ned techniques to handle [ 11 C]carbon monoxide at a nanomole scale. Facilitating biological research and molecular imaging was the driving force when [ 11 C]carbon monoxide was used in the ﬁ rst in vivo application with carbon-11 in human (1945) and when [ 11 C]carbon monoxide was used for the ﬁ rst time as a chemical reagent in the synthesis of [ 11 C]phosgene (1978). This review examines a rich plethora of labelled compounds synthesized from [ 11 C]carbon monoxide, their chemistryand use inmolecular imaging.While thestrongdevelopment ofthe 11 C-carbonylation chemistry has expanded the carbon-11 domain considerably


Introduction
Positron emission tomography (PET) is a non-invasive and quantitative imaging modality used in clinical diagnosis and biomedical research.The technology is based on detection of 511 keV photons emitted from compounds labelled with short-lived positron emitting radionuclides.Carbon-11 with a physical half-life of 20.4 min has the great advantage being naturally occurring in most endogenous and pharmacologically active compounds, hence incorporation of carbon-11 can often be accomplished without compromising the chemical structure.In addition, a number of different strategies can be employed with primary and secondary labelled precursors such as [ 11   [1,2].This review article centres at 11 C-carbonylation methods and the pharmacologically interesting compounds that have been synthesized from [ 11 C]CO.The aim is to give the reader an overview of the field and complement previous reviews covering 11 C-carbonylation [1, [3][4][5][6][7][8][9][10][11].
[ 11 C]CO was not conceived as a labelled precursor in the beginning, but as a tracer to study interactions with red blood cells in vivo.The study was performed in 1945 and was the first human application of a 11 C-labelled compound [12].The first synthesis with [ 11 C]CO came in 1978 with two methods to produce [ 11 C]phosgene, a gas-phase reaction with chlorine gas initiated by photoirradiation [13] and a heterogenous reaction over heated platinum tetrachloride [14].These reports were followed by applications of [ 11 C]phosgene chemistry [15,16] including the synthesis of [ 11   C]methane that gave higher radiochemical yields and molar activities [20][21][22].Although [ 11 C]phosgene was proving its value as a 11 C-carbonyl synthon it could only label a small portion of the rich family of carbonyl compounds.Its scope was mainly labelling of symmetrical ureas, ureas formed via ring closures and carbamates where the high reactivity of [ 11 C]phosgene towards nitrogen nucleophiles did not lead to poor product selectivity.Transition metal catalysed carbonylation with carbon monoxide developed rapidly in the 1970s and made possible to synthesize a wide array of carbonyl compounds from CO, today arguably the most versatile C1 building block for organic synthesis [23].These developments and the fact that the carbonyl group is present in so many biomolecules sparked interest in 11 C-carbonylation protocols and potential production of PET-tracers beyond the limitations in [ 11 C]phosgene chemistry.
While evident that 11 C-carbonylation emerged through the translation of advancements made in organic synthesis, reaction conditions had to be optimized focusing on efficient incorporation of [ 11 C]CO and not on the consumption of substrates and catalyst turn-over numbers.In addition, the production of [ 11 C]CO and techniques to transfer it efficiently to low volume reaction vessels had to be researched.Once these obstacles were overcome, 11 C-carbonylation began to make its way into PET tracer synthesis.However, in retrospect one can argue that a rather limited share of publications involving 11 C-carbonylations have included biological evaluations and naturally even fewer clinical PET applications.Most publications on 11 C-carbonylatons have been chemistry-oriented and focused on method development and mapping out the scope of the new methodologies.The reasons for a rather limited implementation of 11 C-carbonylation protocols in routine PET-tracer production could be a consequence of technical challenges or lack of commercially available synthesis systems.But considering the progress made by relatively few research groups with capability of [ 11 C]CO chemistry, there should be great potential for future growth in this area.

Production of [ 11 C]CO
Carbon-11 is today exclusively produced by the 14 N(p,α) 11 C nuclear reaction, typically using a cyclotron that generates [16][17][18] MeV protons that irradiate a nitrogen gas target mixed with hydrogen or oxygen gas to produce [ 11 C]methane or [ 11 C]carbon dioxide, respectively.[ 11 C]Carbon dioxide is the most versatile in-target produced labelled precursor and can readily be converted to [ 11 C]CO, Table 1.Typically the oxygen content in the target gas varies from 50 ppm to 2% depending on type of target holder.Target holders with relatively small volumes, operated under high target pressures, are typical for more recently developed systems and the oxygen content in the target gas has been increased to optimize recovery of [ 11 C]CO 2 .Approximately 100 GBq [ 11 C]CO 2 can be produced with a 30 min irradiation by a modern cyclotron.Low energy cyclotrons, 8-12 MeV proton energies, can also be used but may render lower radioactivity yields and molar activities, which could be unsuitable for labelling of receptor ligands.It is possible to produce [ 11 C]CO directly in the target holder at low beam current or with a target high in oxygen content, but today [ 11 C]CO is normally produced from [ 11 C]CO 2 via a reduction process [24].Practical on-line methods for the conversion of [ 11 C]CO 2 to [ 11 C]CO have been developed based on heated gas-flow reactors typically filled with either zinc or molybdenum.
Zinc-mediated reduction is typically carried out at 400 °C and should not approach the melting point of the metal, 419.5 °C, or else the granulated zinc loses its reactivity, possibly due to changes in surface area properties [25].When using [ 11 C]CO 2 produced in a low-pressure target (4 bar) with low concentration of oxygen (0.05%), the conversion of [ 11 C]CO 2 to [ 11 C]CO was approximately 80% (n = 3) [26].Essentially all activity not recovered as [ 11 C]CO is typically trapped on the zincmaterial.After some time in use, the reducing activity of the zinc is eventually lost, even when operated at optimal temperature.This can occur abruptly and after varying number of [ 11 C]CO production runs.However, with [ 11 C]CO 2 produced in high-pressure nitrogen gas targets (20-30 bar filling pressure) and oxygen concentrations of 0.5-2%, the depletion of the zinc usually occurs immediately.This can be circumvented by purifying the [ 11 C]CO 2 before the reduction-step as shown by Verbeek et al. [27] The gas purification was performed over a silica gel column and the study indicated that O 2 from the target gas was not a direct cause for zinc-depletion.Instead it was stipulated that the cause was N 2 O, formed in the target holder during the irradiation.Molybdenum reductant (at 850 °C) appear to be more robust compared to zinc but in general the conversion to [ 11 C]CO is lower, around 70% [28,29].Charcoal heated at 800-900 °C also reduces [ 11 C]CO 2 to [ 11 C] CO but the method has not gained wide spread use, potentially due to the risk of isotopic dilution [30][31][32].
Several other methods have also been investigated aiming for robust and high yielding production of [ 11 C]CO regardless of target system.Chemical conversion of [ 11 C]CO 2 via [ 11 C]formate and [ 11 C]formyl chloride to [ 11 C]CO was presented by Roeda et al, however so far the method has not been replicated in the literature [33].Production of [ 11 C]CO has also been done by [ 11 C]CO 2 fixation with silyllithium [34] or disilane [35] in THF, followed by release of [ 11 C]CO through addition of a fluoride source.The reported yields were 74% and 99%, respectively, using nonautomated processes [34,36].The use of these reagent solutions could potentially be a source for isotopic dilution by CO 2 -capture upon storage and handling leading.However, it was found that a stock solution of the silyllithium reagent, prepared from chloro(tert-butyl)diphenylsilane and lithium in tetrahydrofuran could be stored for one week in the freezer with retained reactivity and still be used to synthesize products with molar activities in the range of 300-500 GBq/μmol [34].
A proof-of-principle study on novel electrochemical [ 11 C]CO 2 reduction to [ 11 C]CO was also recently presented [37].The [ 11 C]CO 2 was passed through a solution of aqueous potassium chloride, base and a catalyst (Ni(cyclam) 2+ or Zn(cyclen) 2+ ), with up to 66% trapping of the activity.The reduction was carried out using electrolysis at an electrode potential of −1.8 V.The formed [ 11 C]CO was passed through a solution of Pd-Xantphos, benzyl amine and phenyl iodide at room temperature whereby [carbonyl- 11 C]N-benzylbenzamide was obtained in 1-10% radiochemical yield based [ 11 C]CO 2 .
Early on, Kilbourne et al had used heated zinc supported on asbestos for reduction of [ 11 C]CO 2 to [ 11 C]CO.[38] This concept was revisited by Dahl et al, obtaining an average [ 11 C]CO yield of 93% (n = 20) by the reduction over zinc granules mixed with silica gel heated at 485 °C, i.e. well above the melting point of the metal [39].A very good result considering that the conventional reduction method with only zinc had failed to produce any [ 11 C]CO at all, possibly due to the high oxygen content (0.5%) and formation of N 2 O in the target holder, as discussed previously.While the molar activity was low (11 GBq/μmol) it was deemed normal for the target system used.The method was recently applied by Wildt et al albeit no data on the radiochemical yield of the produced [ 11 C]CO was given [40,41].The molar activity for N-[ 11 C] acylsulfonamide synthesized from [ 11 C]CO was rather low (21 GBq/ μmol), but again deemed normal due to small scale radionuclide production.Therefore, more studies are needed to evaluate its general suitability in PET-tracer synthesis.]cyanides transferred in carrier gas can be captured in reaction mixtures simply by bubbling through at room temperature or lower temperatures.However, in the case of [ 11 C]CO, the poor solubility in most solvents requires special arrangements to accomplish high trapping efficiency in small volume reaction vessels.

Pioneering work
The first description of a carbonylation reaction with [ 11 C]CO dates back to 1983 when Kilbourne et al bubbled [ 11 C]CO in a stream of helium into a cold (−78 °C) solution of lithium piperidide in tetrahydrofuran/ dimethoxyethane [38].Between 10% and 20% of the activity was trapped in the vessel, presumably in the form of [ 11   C]CO through the reaction mixture improved the trapping.The method worked and [ 11 C]ketones were isolated in radiochemical yields of 26-62% [43].However, the modest trapping of [ 11 C] CO with single-pass transfer and the technically cumbersome recirculation method made the methodologies unsuitable for routine use.But these pioneering attempts nevertheless proved that the carbonylative Stille coupling protocol worked using [ 11 C]CO as the limiting reactant.
Many examples of transition metal-mediated 11 C-carbonylations have since followed, often translated from conventional bench-scale catalysis.It should be noted that in 11 C-carbonylation the yield is calculated based on [ 11 C]CO, while in conventional catalysis the yield is based on other more valuable reactants than CO used in excess.Hence, for these two applications of carbonylation chemistry the yield describes two different reaction characteristics.
The high molar activity of [ 11 C]CO means that when GBq amounts of activity is used, the amount of [ 11 C]CO is only a few nanomoles.Thus, [ 11 C]CO is employed in extreme substoichiometric amounts to both transition metal complexes and the substrates (micromoles), often facilitating fast consumption of [ 11 C]CO and render the propagation of the catalytic cycle redundant or in fact unlikely.The use of low amounts of [ 11 C]CO can also present challenges as conventional carbonylation may not always be readily reproduced in 11 C-labelling, for example in reactions where CO simultaneously plays other roles besides being a substrate, such as a reductant or a spectator ligand.

High-pressure autoclave system
Improved techniques to confine [ 11 C]CO in reaction vessels were needed to accomplish efficient 11 C-carbonylation protocols.This was addressed by the high-pressure autoclave system, Fig. 1 [44].[ 11C]CO 2 was transferred in nitrogen gas to the system where it was trapped and concentrated on a silica column at liquid nitrogen temperature.The trap was then heated to release the [ 11 C]CO 2 in a stream of helium gas that was passed through a reductant (zinc at 400 °C) and then through an Ascarite column to remove traces of remaining [ 11 C]CO 2 .The formed [ 11 C]CO was concentrated on a small silica column (<10 μL) submerged in liquid nitrogen.It was essential to exchange the nitrogen transfer gas to helium to quantitatively trap [ 11 C]CO on the small silica column.A valve vas then switched to block the line between the trap and the micro-autoclave (200 μL) giving rise to a pressure build-up of approximately 400-500 kPa by the helium carrier gas.The [ 11 C]CO was then released from the silica material by heating the trap and subsequently transferred to the micro-autoclave by the pressure equilibration upon opening the valve.After the transfer of [ 11 C]CO to the autoclave the valve was switched to stop again.The reagents were then transferred from the reagent injection loop to the micro-autoclave by the solvent pumped at 41 MPa.This resulted in a reduction of the gas-phase volume in the autoclave to <3 μL.High recovery of [ 11 C]CO in the autoclave relied on efficient mass transfer achieved by significantly smaller volume in the CO-trap and transfer tubing compared to the autoclave.
The autoclave method was essential for the development of 11 C-carbonylation chemistry and opened for production of PET tracers with high radioactivity yields.The fact that so many compounds labelled by 11 C-carbonylation has been synthesized using this technology, >490 published compounds and 60 research articles, clearly shows its value as a research tool.However, due to the technical complexity of the highpressure system and lack of commercially available modules, only few PET sites have implemented this method for GMP production and a handful of clinical tracers have been prepared.
In addition, the re-use of the same reactor for all productions makes it challenging to validate that carry-over of reagents or by-products from earlier productions is not occurring.If the autoclave is not cleaned sufficiently to remove residues of for example transition metals, active in very small concentrations, even repeated runs of the same synthesis could be negatively influenced.For example, during productions of the glycine antagonist [ 11 C]GV15026 41, Scheme 1, an increase of trans-to-cis isomerization occurred in the reactions (previously not published data).
The trans-configuration was expected to be preserved during the reaction, and this proved to be the case during the first syntheses.However, after extended use of the autoclave with Pd(0)-catalysts, the corresponding trans isomer ([ 11 C]GV170651) increasingly appeared and the ratio between trans and cis gradually decreased from 95/5 to approximately 60/40.This may have happened due to deposition of Pd(II) formed from Pd(0), on the stainless-steel surface of the autoclave during the reactions.The Pd(II) could catalyse the isomerisation via formation of a π-Pd-complex with the vinyl iodide precursor.For this reason the standard cleaning procedure for the stainless-steel autoclave was changed to also include wash with aqueous nitric acid (10%) to remove Pd(II).After the introduction of the acid wash the E/Z ratio of 95/5 was regained, clearly stressing the importance of thorough cleaning procedures.A cleaning and conditioning process to reduce the carryover effect between syntheses in autoclaves for 11 C-carbonylation has been proposed by Karimi et al describing the resulting reproducibility of 11 C-aminocarbonylations to be higher than 95% [46].

Confining [ 11 C]CO at low pressure
Following the autoclave method, several methods have been researched to facilitate carbonylation reactions at low pressure, e.g. by chemical complexation, transfer in soluble carrier gas, transfer to vacuumized vials and ex situ production of  with the aim to confine [ 11 C]CO together with the carbonylation reagents in small volume glass vessels.

Trapping by boron complex
Complexation with BH 3 •THF was the first example where an additive was used to enhance the trapping efficiency of [ 11 C]CO (>90%) [47].The method required low trapping temperature, −78 °C, and the reaction solvent was limited to THF.The radiochemical yield of [ 11 C]Nbenzyl benzamide was low but improved to 47% when water (1%) was added to release the [ 11 C]CO from the BH 3 •[ 11 C]CO complex.No further applications have been reported since the original publication.

Trapping by copper complexes
Kealey et al discovered that [ 11 C]CO could be trapped at ambient pressure by the copper (I) tris(pyrazolol)borate complex, named Scorpionate or Cu[Tp*] and then be released by addition of triphenylphosphine, Scheme 2 [48].The trapping efficiency of [ 11 C]CO was increased from <1% to 96% when adding the Cu[Tp*] complex to pure THF and 99% of the trapped [ 11 C]CO was recovered after the release.The additive did not perturbate the carbonylation reactions mediated by Pd(dppe)Cl 2 or Pd(dba) 2 yielding [ 11 C]N-benzylbenzamide (67%) and [ 11 C]dibenzylurea (47%).Later it was discovered that [ 11 C] CO could be released by the phosphine ligands present in the precatalyst complex, e.g.Pd(PPh 3 ) 4 or Pd(PPh 3 ) 2 Cl 2 [49,50].The Cu[Tp*] complex was used in syntheses of [ 11 C]amides [49,51,52], [ 11 C]ureas [50] and a lactam [53].The synthesis was also implemented on a microfluidic device [52,53].The complex has not yet been tested with other reaction mediators than palladium, e.g.rhodium or radical initiators.A second generation of tris(pyrazolyl)borate ligands has also been investigated, with focus on interactions between CuI and [ 11 C]CO/CO.[54] Their practical use for [ 11 C]CO capture and release was not improved compared with the original Cu[Tp*] complex.

Trapping by iron, cobalt and nickel complexes
Three Fe(II) complexes have been explored with the aim to trap [ 11 C] CO in solution and further release it upon irradiation with UV-light [55].All three systems worked well with CO, however, none of them was successfully translated to microscale [ 11 C]CO chemistry.Two of the ligands could not efficiently trap [ 11 C]CO and the third, (TPP)Fe(THF) x showed high trapping efficiency (97%) but failed to release [ 11 C]CO upon UV irradiation.
A novel P-S-N ligand forming Fe(II), Co(II) and Ni(II)-complexes showed ability to trap CO (at 1 atm.) and release it upon heating [56].However, [ 11 C]CO could not be trapped, likely due to slow carbonyl adduct formation.

Trapping by palladium-Xantphos complexes
Xantphos was introduced to Pd-mediated 11 C-carbonylation by Miller et al as an good support ligand for labelling amides and a carboxylic ester using a microfluidic device [52,57].Dahl et al discovered that the high reactivity of the Pd-Xantphos complex enabled near quantitative trapping of [ 11 C]CO when it was bubbled into the 4 mL reaction vial [58].Besides producing a series of N-benzyl-[carbonyl-11 C]benzamides the scope of the Pd-Xantphos complex was tested in the synthesis of an [ 11 C]aldehyde, a [ 11 C]ketone and a [ 11 C]carboxylic acid, also obtained with high radiochemical yields (54-78%).A histamine type-3 receptor ligand, later named [ 11 C]AZ13198083 51, was synthesized in 95% radiochemical yield by 11 C-aminocarbonylation. Hence, it was concluded that the Pd(0)-complex supported both a range of carbonylative coupling processes and efficient trapping of [ 11 C]CO, essentially removing the need for chemical-trapping additives or technically advanced equipment.
However, there might be a potential pitfall associated with the Xantphos ligand.A significant [ 11 C]benzamide side-product (36-51%) appeared in the synthesis of [ 11 C]Olaparib 25 (<1%) which likely derived from a phenyl group transferred from the Xantphos ligand to the Pd metal centre [59].This observation underlines the importance of careful analysis of the radiolabelled product, especially when using derivates of aryl halides as substrates in the carbonylation reactions, given the risk for erroneous characterisation of the product due to inadequate separation from the side-product by analytical HPLC.The same sideproduct was also obtained with triphenylphosphine as supporting ligand, albeit in lower amount (4%).

Transfer of [ 11 C]CO in xenon gas to sealed reaction vial
A method to confine [ 11 C]CO in small-volume disposable glass vials at low pressure has been developed by Eriksson et al. [60,61] The method exploited the remarkable solubility of xenon in THF to achieve nearly quantitative transfer of [ 11 C]CO into non-vented reaction vessels without significant pressure build-up.Since the vials lacked venting the transfer gas remained inside, thus preventing [ 11 C]CO from escaping.While using the same techniques to concentrate [ 11 C]CO 2 and [ 11 C]CO on cold-traps as in the high-pressure autoclave system [44], the method was technically less complicated.The process required only one multiport valve for directing gasses, and the fact that no solutions were handled by the valve made it practically maintenance free, Fig. 2. Furthermore, the use of disposable reaction vials, standard vials for HPLC applications (1 mL), reduced the risk for carry-over from previous syntheses.Vacuumized vials could also be used, e.g. for efficient transfer of [ 11 C]CO if the selected solvent does not dissolve xenon gas well.The 'xenon-method' has been applied in transition metal mediated synthesis of [ 11 C]amides [60,62,63], [ 11 C]ureas [60,64], [ 11 C]carboxylic esters [60], [ 11 C]sulfonyl carbamate esters [26], [ 11 C]ketones [65] and N-[ 11 C] acylsulfonamide [66].These results indicate that the radiochemical yields for transition-metal mediated reactions are not dependent on solvent-gas phase volume ratios, which could have justified use of high-pressure autoclaves.However, such correlation favouring higher solvent-gas phase ratios was observed for thermally initiated radical 11 C-carbonylation of aliphatic alkyl iodides, i.e. in reactions without transition metals [61].Another advantage using an autoclave is the possibility to heat reactions at very high temperatures.

Transfer of [ 11 C]CO in helium gas to sealed reaction vial(s)
Wildt et al recently devised a dispensing system that concentrated [ 11 C]CO on a silica-trap submerged in liquid nitrogen, which after release enabled transfer of the [ 11 C]CO in helium to a motorised syringe from where it was dispensed into multiple vials sealed with septa [40].Similar to the 'xenon-method' the [ 11 C]CO was prevented from escaping by not venting the vials during the transfer [60,61].The recovery of [ 11 C] CO in the vials was moderate (66%) and the activity was unevenly distributed due to pressure build-up, nevertheless, synthesis of [ 11 C]Nbenzylbenzamide in up to 86% radiochemical yield proved the usefulness of the method.Additionally, a protocol was outlined where ten products were synthesized in parallel and analysed simultaneously by radio-TLC.The [ 11 C]CO dispensing method was later used in the synthesis of a series of N-[ 11 C]acylsulfonamides [41].

"In-loop" 11 C-carbonylation
A new technique for 11 C-carbonylation reactions was recently presented by Ferrat et al that utilized a stainless-steel loop as reactor, connected to an HPLC injection valve [67].The method allowed 11 C-carbonylation to be performed analogue to the technique developed by Wilson et al for 11 C-methylations [68].Reagents were loaded in the loop prior transfer of [ 11 C]CO in helium gas.Quantitative trapping of [ 11 C]CO was achieved in the loop in all reactions using metal-complex prepared from Pd 2 [π-cinnamyl]Cl 2 and Xantphos, and also when the ligand was switched from Xantphos to P t Bu 3 , indicating that the method has a wider scope.A set of seven compounds were reported with impressive radiochemical yields, including [ 11  AZ13198083 51 (91%).The latter compound was also purified and reformulated and was then obtained in 38% radiochemical yield.
While the method requires preconcentration of [ 11 C]CO like many other 11 C-carbonylation techniques, the overall complexity should be sufficiently low to be an attractive alternative, especially since other "in-loop"-reactions have been used successfully in production of clinical PET-tracers.

Flow chemistry and microfluidic systems
The first example of 11 C-carbonylation performed using flow chemistry was presented by Miller et al. [69] [ 11 C]CO was passed through a reactor consisting of Teflon tubing (45 cm in length and 1 mm in diameter) packed with a Pd-phosphine complex immobilized on silica.The reactor, preloaded with aryl halide and benzyl amine, was heated at 80 °C for 6 min where after the formed [ 11 C]amide was eluted by solvent during 5 min.The reactor was reused a second time when repeating the same synthesis but was exchanged when changing to different substrates.
In 2011 two papers on micro-scale reactors for 11 C-carbonylations were published.Miller et al reported the synthesis of [carbonyl- 11 C]   phthalide via intramolecular alkoxycarbonylation (87%) and a series of 11 C-labelled benzylamides (44-88%) via aminocarbonylation of aromatic iodides, this time using a glass-fabricated micro-channel reactor operating in the annular flow mode [52].[ 11 C]CO was introduced in a stream of nitrogen gas and a syringe pump pushed the reaction solution through the microfluidic device containing channels (5 m in length and 220 × 100 μm wide).After the 10 min reaction time the device was flushed with 100 μm solvent to collect the product.
Kealey et al used a commercial device (NanoTek, Advion) to perform microfluidic 11 C-carbonylations in a silica capillary.First [ 11 C]CO was pre-trapped in a solution of Cu[Tp*] and subsequently this solution was transferred into the microreactor simultaneously with a second solution containing all other coupling reactants [53].[ 11 C]MK-0233 43 was labelled in this way at the γ-butyrolactone carbonyl position in 7% radiochemical yield.The reaction returned similar result when performed in a glass vial.
Recently, a segmented-flow microfluidic method was presented by Dahl et al. [70] [ 11 C]CO in helium carrier gas was introduced together with a reagent solution into a silica capillary (5 m in length and 200 μm in diameter) via a mixing tee resulting in a flow of alternating liquid and gas segments inside the capillary, which was heated at 100 °C.In total 12 model compounds were synthesized, three esters (79-99%), one carboxylic acid (95%) and eight amides including D 2 receptor radioligands [ 11 C]FLB457 30 (61%) and [carbonyl- 11 C]Raclopride 55 (79%).[42].Several papers on ketone labelling followed using more efficient techniques to confine and react the [ 11 C] CO with aryl iodides [30,31,43,58,[72][73][74], vinyl triflate [43], methyl iodide [43,73] and diaryliodonium bromide and triflate [30].Typically, methyl and phenyl stannanes were used as transmetalation reagents.Karimi et al expanded the scope with e.g.pyridine-, thiophene-, furan-and higher alkyl-stannanes as well a few of the corresponding organoiodides [73].Many of the products were obtained in radiochemical yields ranging between 40 and 90%.Slobbe et al utilized Stille coupling to label a mutation selective serine/threonine kinase inhibitor, [ 11 C]Vemurafenib 59 (21%), Scheme 3 [65].The compound was evaluated in mice melanoma xenografts by autoradiography, biodistribution and metabolite analysis with the aim to develop a method for identifying tumours sensitive to vemurafenib treatment.The compound had similar uptake in Colo829 (BRAFV600E) and MeWo (BRAFwt) xenografts indicating difficulties to distinguish treatment responding BRAFV600E mutations from BRAFwt in vivo using [ 11 C]vemurafenib PET.To the best of our knowledge, the study by Slobbe et al was the first imaging application with a ketone labelled by 11 C-carbonylation, despite published as late as 2017.Siméon et al synthesized four tracer candidates for imaging of presynaptic histamine subtype-3 receptors 60 (4.9-8.6%).The Pd(0)-mediated carbonylation of aryl iodides and aryltrimethylstannanes was performed under anhydrous reaction conditions to suppress formation of labelled carboxylic acids [74].

Carbonylation with [ 11 C]CO
In 1997, Zeisler et al presented the first Suzuki carbonylative coupling with [ 11 C]CO and introduced heated molybdenum as a reductant to produce [ 11 C]CO from [ 11 C]CO 2 [29].Nader et al used the method to synthesize a weak inhibitor of human immunodeficiency virus type 1 (HIV 1) reverse transcriptase 5 (71%) by reacting phenylboronic acid with an iodoaryl compound and [ 11 C]CO in the presence of Pd (PPh 3 ) 2 Cl 2 and potassium carbonate, Scheme 4 [72].The trapping efficiency of [ 11 C]CO (36-40%) was higher compared to the Heck reaction (approximately 10%) [42].These two pioneering papers on Suzuki coupling used Pd(II) complexes while later studies have used Pd(0).The generally accepted mechanism for the Suzuki coupling proceeds via Pd (0) and thus it can be assumed that the Pd(II) was reduced in situ, possibly also partly by the oxidation of [ 11 C]CO, which may explain the high trapping efficiency.
Rahman et al studied the influence of different bases on the radiochemical yields of [ 11 C]ketones (14-74%) prepared by Pd(0)-mediated Suzuki coupling [75].Best results were obtained with tetrabutylammonium fluoride or potassium tert-butoxide base.However, contrary to common observations that base must be added, supposedly to activate the boronic acid, in two cases the radiochemical yields were better without added base.In a related study, potassium fluoride, which served as a base caused blockage in the high-pressure carbonylation system and therefore was removed from the synthesis protocol.The syntheses worked equally well without potassium fluoride and fifteen compounds were obtained in 10-64% isolated radiochemical yield [76].It could be speculated that base is not needed in Suzuki type

11 C-Aminocarbonylation
The first paper on Pd-mediated aminocarbonylation was published in 1999 by Kihlberg et al and included carbonylation of benzyl bromides and aryl iodides with 20-85% and 43-76% radiochemical yields, respectively [77].Four secondary aliphatic amines and one primary amine were synthesized in addition to [ 11 C]Benzotript 3, a cholecystokinin receptor antagonist obtained in 55% radiochemical yield, Scheme 5.

11
C-Aminocarbonylation mediated by Pd(0) can mechanistically be described as a three-step sequence starting with oxidative addition of an organohalide/triflate lacking β-hydrogens, Scheme 6.The obtained organo-Pd complex is active towards [ 11 C]CO insertion and formation of a Pd-[ 11 C]acyl complex which then reacts with an amine to produce the labelled amide via reductive elimination.The oxidative addition of organoiodides is not rate determining in 11 C-carbonylation, as it is for conventional carbonylation with CO (isotopically unmodified).The catalytic cycle does not propagate during the reaction due the large excess of organo-Pd complex compared to [ 11 C]CO, in fact, sufficient concentration of organo-Pd complex may already have formed in the time between the preparation of the reaction mixture and the addition of [ 11 C]CO.This may not be the case for less reactive substrates, e.g.organobromides, where preheating the Pd-complex and organohalide mixture prior to addition of [ 11 C]CO have shown to improve the radiochemical yields [46,52,73,[78][79][80][81][82].The transformation to the new Pdcomplex is often accompanied by a change in colour tone of the reagent solution.Successful aminocarbonylation of organochlorides have also been reported, accomplished by pre-heating [79], rapid microwave heating during the reaction [83] and elevated reaction temperature (170 °C) [81], respectively.
The third component in aminocarbonylation is the amine which acts as nucleophile.Reactions with less nucleophilic amines may benefit from increased reaction temperatures and amine concentrations.Activation of less reactive amines by strong base was often useful to improve the radiochemical yield, e.g. by butyl lithium [84,85], pempidine [79], lithium bis(trimethylsilyl)amide (LiHMDS) [46,62,79] and 7-methyl-1,5,7-triazabicyclo[4.4.0]dec-5-ene (MTBD) [59].It should be noted that base plays a role in conventional Pd-mediated carbonylation to remove the acid formed by the oxidative addition, this may be less important in the case of 11 C-carbonylation as substoichiometric amounts of [ 11 C]CO prevents the catalytic process to propagate. 11C-Labelled primary amides have been synthesized from aryl iodides and iodo/ bromopyridines with ammonia by Pd-mediated carbonylation at high temperatures (180 °C), e.g.nicotinamide 7 (11-54%), Scheme 7 [78].As shown in the early work of Kilbourne et al and later by Itsenko et al, [ 11 C]formamides can be synthesized from [ 11 C]CO by direct reaction with lithium amides, without use of transition metals, exemplified with lithium 1-phenylpiperazide (75%) [38,85].
Up till now, close to 170 [ 11 C]amides have been synthesized using Pd-PPh 3 -complexes, overshadowing the approximately 60 compounds obtained by Pd-Xantphos-complexes.However, counting from 2013 when it was known that Xantphos could efficiently trap [ 11 C]CO, the numbers are 45 to 16 in favour to Xantphos.The radiochemical yields are often high with Pd-Xantphos-complexes suggesting that the bidentate supporting ligand may be more efficient than PPh 3 for many applications.The solvent of choice has predominantly been THF while DMF have been beneficial in some cases.THF was also the preferred solvent for microwave heating when screening solvents for the synthesis of 3-nitrobenzyl-[carbonyl- 11 C]benzamide [83].
A series of [ 11 C]N-cyanobenzamides were synthesized by Pd(0)-mediated aminocarbonylation with 1,1′-bis(diphenyl-phosphino) ferrocene (dppf) as a support ligand [81].The reaction scope was studied by labelling of nine compounds using different aryl iodides (28-79%) and nine compounds using aryl bromides (34-72%).The optimized reaction conditions were used to synthesize biologically interesting 11  While most publications involving 11 C-aminocarbonylations have focused on the development of labelling methods, a number of publications also include preclinical evaluation of synthesized [ 11 C]amides.The earliest example was a [ 11 C]Citalopram analogue 42 (62%) obtained by aminocarbonylation of an aryl bromide, published by Madsen et al. [96] Biodistribution in rat showed very low penetration of the bloodbrain barrier (BBB) and the labelled amide was hypothesized to be a P-gp substrate.Åberg et al evaluated angiotensin II AT 2 receptor ligands 47 (16-36%) with the 11 C-label incorporated using benzyl-, diethyl-and isopropylamine via aminocarbonylation of aryl iodide [93].The isopropyl analogue was subjected to biological investigation due to its favourable lower lipophilicity.In vitro autoradiography on frozen sections revealed selective binding in rat pancreas and kidney cortex and to some degree in pig adrenals and rat brain.Biodistribution in rat showed high uptake in excretory organs and rapid accumulation in urine, observed with PET imaging in rat, suggested low metabolic stability.Nordeman et al labelled [ 11 C]BSI-IV 52 via aminocarbonylation using both aryl iodide and aryl bromide [89].The radiochemical yields were 29% and 7%, respectively.Autoradiography showed low specific binding.Ex vivo biodistribution and PET imaging in rat showed fast clearance and low brain uptake indicating that the compound had limited use in the study of Alzheimer's disease or BACE-1.Bergman et al labelled six novel Benzovesamicol derivatives 53 in 5-25% radiochemical yield with the aim to image the vesicular acetylcholine transporter system (VAChT) [88].Two compounds showed specific VAChT binding in human post mortem autoradiography studies on brain tissue from Alzheimer's disease patients, but not high enough to justify further preclinical studies.Recently, a second paper was published focusing on syn-thesis of isotopically unmodified compounds and evaluation of their affinity and selectivity for VAChT, one compound was 11 C-labelled 35 (10%) [98].Hong et al synthesized [ 11 C]FIMX 54 a metabotropic glutamate receptor 1 (mGluR1) ligand obtained in 5% radiochemical yield via carbonylative coupling of aryl halide and amine, followed by BOCdeprotection [90].The compound was used in Rhesus monkey PETimaging and the uptake observed in the base-line scan was reduced after receptor blocking which indicated selectivity for the target.Metabolite analysis showed 21% intact tracer after 30 min and 4 metabolites were detected.Rahman et al synthesized [carbonyl- 11 C]Raclopride 55 in 50% radiochemical yield, an isotopomer of the well-established dopamine D 2 ligand [methyl- 11 C]Raclopride, Scheme 9 [91].The compounds were compared in cynomolgus monkeys and showed similar results.As the two compounds were structurally identical but with the label at different positions, metabolic transformation in vivo could have resulted in differences between the two tracers.However, metabolite analysis revealed similar metabolite patterns and the authors concluded that metabolic cleavage did not occur at the methoxy group.
Van der Wildt et al labelled acrylamides by aminocarbonylation, 56 and 57, three candidates for irreversible binding to tissue transglutaminase (TG2) [62].The compounds were obtained in 38-55% radiochemical yield.Their biodistribution and metabolic profile were evaluated in rat and the most metabolically stabile compound was further evaluated by autoradiography on tumour tissue which showed selectivity to the active state of TG2.
Recently Dahl et al synthesized three compounds targeting the histamine type-3 receptor (H 3 R) involved in release of the neurotransmitters histamine, dopamine, and noradrenaline [99].All compounds were labelled using the same 11  molgus monkey showed that all three compounds crossed the bloodbrain barrier and had brain distribution consistent with H3R expression.Pre-treatment and displacement studies with AZD5213 showed that [ 11 C]AZ13198083 57 had high specific uptake and rapid reversible binding.
Hydroxycarbonylation is typically performed in organic solvents with addition of water and a base, such as trimethylphenylammonium hydroxide [80], tetra-N-butylammonium hydroxide [80,101], tetra-Nethylammonium hydroxide [28,58] and sodium hydroxide [70,83].Ishii et al synthesized [ 11 C]aspirin 31 (12%) by carbonylation of a boronic ester in base-free DMF-water, noting that the yield was not improved by addition of tetrabutylammonium hydroxide [102].Åberg et al synthesized [ 11 C]Eprosartan 46 (37-54%) by hydroxycarbonylation of aryl iodide in the presence of water and tetra-N-butylammonium hydroxide, Scheme 10 [101].Both papers suggest the need to fine-tune the reaction conditions to get high radiochemical yields and avoid technical issues such as precipitation.
[ 11 C]Eprosartan 46 was synthesized with the aim to develop a highaffinity AT 2 -receptor PET tracer and was evaluated by frozen-section autoradiography in rat and pig tissue which showed specific binding in kidney, lung and adrenal cortex.A biodistribution study in rat revealed high accumulation in kidney, liver and intestinal wall.

11 C-Alkoxycarbonylation
Heck and co-workers described the first Pd(0)-catalysed alkoxycarbonylation reaction in 1974 [103].The strength in carbonylative coupling reactions lays in the large variety of products that can be prepared by joining two substrates with the carbonyl functionality, often under mild conditions acceptable to many types of functional groups.Considering the importance of [ 11 C]methyl iodide for PET, where the incorporation of the labelled methyl group has become a workhorse for 11 C-tracer production, it is easy to understand that well-developed carbonylation protocols could play an important part to expand the diversity of PET chemistry.Alkoxycarbonylation mediated by Pd(0) gives direct access to carboxylic esters in one step, however obviously limited to substrates lacking β-hydrogens, e.g.aryl iodides [70,83,104], methyl iodide [105] and iodopyridine [87].Several instances of intramolecular coupling reactions forming lactones are found in the literature, mostly serving as convenient model reactions as the same substrate contains both the alcohol group and an aryl iodide [28,47,52,58,70,106] or aryl bromide [53,92].In addition to organohalides as substrates, Ishii et al labelled twenty different methyl esters (6-80%) by low temperature carbonylation of heteroaryl boronic acid pinacol esters in the presence of methanol, Pd(OAc) 2 , triphenylphosphine and p-benzoquinone [102].A few examples were also shown of labelled esters treated with sodium hydroxide or aqueous ammonium to give the corresponding carboxylic acids and amides, respectively.
As indicated by high demand for methylated 11 C-tracers, including methyl esters, several biologically interesting carboxylic esters have also been synthesized using 11   C]Acetate is a potential marker of cerebral oxidative metabolism and glial cell metabolism but suffers from a low brain uptake.The authors concluded that [carbonyl- 11 C]benzyl acetate had a relatively high peak uptake (SUV = 3.1) and may be used to measure uptake and metabolism of acetate in glial cells of the brain.

[ 11 C]Aldehyde synthesis
The first synthesis of [  [109].The formylation reaction proceeded with similar radiochemical yields at temperatures from room temperature up to 70 °C, but above this temperature formation of [ 11 C]carboxylic acid started to hamper the reaction.

[ 11 C]Urea synthesis
Two protocols for the synthesis of [ 11 C]ureas by Pd(II)-mediated oxidative carbonylation have been published [50,64].After observing [ 11  analytical HPLC) [50].The use of equimolar amounts of two different amines in attempts to obtain unsymmetrical ureas gave largely mixtures of labelled ureas with product distribution depending on the amine reactivity.[carbonyl- 11 C]GSK1034702 51 targeting the M1 muscarinic acetylcholine receptor, previously radiolabelled at the methyl position, was obtained in 6% radiochemical yield by intramolecular ring closure [50].Roslin et al synthesized five symmetrical and nine unsymmetrical [ 11 C]urea derivatives utilising the Xenon-method for 11 CO transfer [64].The labelled compounds were isolated with semipreparative HPLC in radiochemical yields up to 65%, including an inhibitor of soluble epoxide hydrolase 39 (41%).Optimized reaction conditions towards unsymmetrical [ 11 C]ureas utilized 6 equivalents of piperidine over benzyl amine and gave excellent product selectivity.
[ 11 C]Ureas could not be obtained with two secondary amines, echoing the findings of Kealey et al. [50] This was further investigated by computational chemistry which showed that isocyanate formation from a secondary amine was unfavourable [64].Control experiments with Pd (0) (Pd(PPh 3 ) 4 ) pre-catalyst gave no desired product, further supporting the Pd(II)-mediated reaction pathway via labelled isocyanates.

[ 11 C]Imide synthesis
Imides were labelled early in the progression of Pd(0)-mediated 11 C-carbonylation.The pioneering paper, and so far the only one, describes the synthesis of three cyclic [ 11 C]imides (45-73%) via carbonylative ring closure of 4-bromobenzamide derivatives, including the racemic form of [ 11 C]Thalidomide 4 (69%), Scheme 14 [110].An open-chain imide (77%) was synthesized from 4bromomethoxybenzene and 2-pyrrolidone at a rather extreme temperature (200 °C) made possible using a stainless steel autoclave.The rationale for the very high temperature may have been the poor nucleophilicity of the pyrrolidone nitrogen.

N-[ 11 C]Acylsulfonamide synthesis
The first labelling of N-acylsulfonylamides, performed by 11 C-carbonylative coupling of phenyl iodides and bromides in the presence of DBU-base was recently published by Wildt et al. [41] After screening Pd-complexes, PdCl 2 (PPh 3 ) 2 with Xantphos supporting ligands was selected for the synthesis of fifteen N-[ 11 C]acylsulfonamides from iodide substrates, including the labelling of two anticancer com-pounds, the candidate Tasisulam and the approved drug ABT-199.[ 11 C] Tasisulam 63 and [ 11 C]ABT-199 64 were obtained in 31% and 84% radiochemical yield, respectively.Another four compounds were obtained from phenyl bromide substrates (57-90%).
Schembri et al developed a Pd-catalysed synthesis of acylsulfonamides via carbonylative coupling of indole and pyrrole nucleophiles with sulfonylazides [66].The reaction did not rely on the use of aryl halides, was compatible with low CO pressures and did not require the addition of base or external palladium ligands.According to the authors, this was the first example of the carbonylation of an azide substrate with a carbon-based nucleophile.The method also worked with [ 11 C]CO when swapping the Pd(II)-precatalyst for Pd 2 (dba) 3 and PPh 3 to avoid side-reactions consuming [ 11 C]CO, 11 C-carbonylation of N-methylindole and tosyl azide proceeded with 10-15% radiochemical yield, Scheme 15.

11 C-Carbonylation of olefins and acetylene
Olefins and acetylenes can be reacted with carbon monoxide in hydroformylation and Reppe type reactions to form aldehydes and carboxylic acid derivatives, respectively.Despite being an important research field with many applications outside PET-chemistry, these chemical transformations have not yet been thoroughly investigated as potential 11 C-labelling methods.

Stepwise Pd-mediated 11 C-carbonylation and multi-step syntheses
In search for more efficient syntheses of [ 11 C]amides by Pd(0)-mediated carbonylation, amines were activated in situ by treatment with lithium bis(trimethylsilyl)amide [46].Using this method most of the 11 Clabelled amides were obtained in high radiochemical yields in one-pot carbonylation protocols but the synthesis with 3-methyl-1H-indole as substrate failed using the one-pot reaction.When performing a twostep synthesis protocol, the radiochemical yield was 32%.Using this protocol a Pd-[ 11 C]acyl complex was first formed in the high-pressure autoclave and then the reaction mixture was added to a solution containing 3-methyl-1H-indole, pre-treated with butyl lithium and trimethyltin chloride.Another study, involving syntheses of eleven [ 11 C]amides from lithium amides, concluded that the two-step method improved the radiochemical yields for unreactive amides [85].Nine of the synthesized compounds were analogues of the 5-HT 1A ligand WAY-100635 14.
Scheme 13.Synthesis of benzohydrazides [78].tions conducted in anhydrous DMF and quenched with water/MeCN, compared to one-step protocols in aqueous reaction conditions.Interestingly, both steps were successfully performed at room temperature.In the same study, three [ 11 C]amides (25-30%) were obtained by quenching with methylamine at 50 °C under anhydrous conditions.
Another [ 11 C]amide synthesized using a similar stepwise protocol was the candidate ligand 62 (6%) targeting the brain serotonin subtype 1B receptor (5-HT 1B ).PET investigation in monkey and rat showed low brain-uptake, with slight increase after P-gp block with Tariquidar.Metabolite analysis monkey plasma showed a favourable profile; however, no specific binding was detected in 5-HT 1B blocking experiments performed in monkey and rat.
Four potential ligands 61 (1-5%) for cytosolic phospholipase A2α (cPLA2α) were labelled by forming Pd-[ 11 C]acyl complexes under anhydrous conditions in THF, followed by quenched with water to obtain the labelled carboxylic acids [111].PET in mice revealed poor blood-brain barrier penetration for all four compounds.
Naturally, 11 C-carbonylation reactions can also produce useful intermediates for multi-step syntheses.[ 11 C]N-benzylacrylamide (51%) was synthesized via [ 11 C]N-benzylacrylic acid, conversion to the [ 11 C]acid chloride and acylation of benzyl amine, Scheme 16 [45].The reason for performing a multiple-step protocol, and not direct carbonylative coupling, was precipitation of the benzylamine salt under the acidic conditions used in acetylene 11 C-carbonylation.
The syntheses of the RAR-γ receptor ligand [ 11 C]Am80 21 (48%) [113], also referred to as [ 11 C]Tamibarotene (16%) [81], was obtained by hydrolysis of the corresponding methyl ester.In another study [ 11 C]methyl esters were converted to labelled primary amides by treatment with aqueous ammonium, and to [ 11 C]carboxylic acids by hydrolysis with sodium hydroxide.[102] Another example of synthesis via [ 11 C]carboxylic ester intermediate is the histone deacetylase 6 ligand 11 C]Tubastatin A 33, labelled in the hydroxamic acid position by Lu et al. [104] The compound was synthesized via two different routes.The indirect route via hydroxyaminolysis of the corresponding phenyl ester (16%) gave higher radiochemical yield, Scheme 17, compared to direct Pd(0)-mediated carbonylation of aryl iodide and hydroxylamine (<5%).
An advantage over the conventional synthesis method with Grignard reagent was the high molar activities attained, ranging from 87 to 270 GBq/μmol depending on alkyl iodide.Syvänen et al labelled two analogues of the NK 1 -receptor ligand GR205171, [O-ethyl- 11 C] GR205171 and [O-propyl- 11 C]GR205171 45, by alkylation with [1- 11 C] ethyl iodide and [1-11 C]propyl iodide [114].PET imaging in rhesus monkey and guinea pig revealed that the propyl analogue had no specific binding in striatum, whereas the ethyl analogue showed reversible binding in contrary to the slow dissociation rate observed for [O-methyl- Dahl et al recently devised a two-step protocol where Pd-mediated carbonylation of aryl iodides produced [ 11 C]benzoyl chlorides and these highly reactive intermediates were then used in acylation reactions to synthesize [ 11 C]amides (80-87%) from amines, [ 11 C]carboxylic ester (79%) from methanol and [ 11 C]primary amide (10%) from ammonia.[ 11 C]Carboxylic acid (93%) was obtained by hydrolysis with sodium hydroxide and [ 11 C]aldehyde (55%) by sodium hydride reduction [116].The radiochemical yields obtained by the two-step protocol indicated great potential for synthesis of PET-tracers with higher radiochemical yields than with previously used one-step methods as demonstrated by the high yielding synthesis of [ 11 C]Tamibarotene 21 (83%).The utility of the protocol was further investigated by Roslin et al by reacting [ 11 C] acid chlorides with metalloid reagents [82].Ten [ 11 C]benzyl alcohols (24%-90%) were obtained by treatment with lithium aluminium hydride or lithium aluminium deuteride, eight [ 11 C]benzaldehydes (65%-91%) by treatment with tributyltin hydride or sodium borohydride and four [ 11 C]phenyl ketones (77%-95%) using sodium tetraphenylborate.In addition, two 11 C-labelled compounds of each product class were synthesized from aryl bromides, at elevated temperatures during the carbonylation step to achieve similar radiochemical Scheme 16.Synthesis of [ 11 C]N-benzylacrylamide via three-step synthesis route [45].

Selenium-mediated 11 C-carbonylation
Selenium reacts with carbon monoxide and primary amines to give ammonium carbamoselenoates, which can be converted to symmetric ureas by aminolysis.Furthermore, it is possible to obtain carbamates and carbonates in reactions with oxygen nucleophiles.Kihlberg et al applied the method with [ 11 C]CO to label six carbamates (22-84%), three ureas (38-88%) and one carbonate compound (78%) [117].Selenium has inadequate solubility in solvents apt for carbonylation, however a soluble selenium-complex is formed with tetrabutylammonium fluoride in DMSO.The method was used to synthesize [carbonyl- 11 C] Zolmitriptan 49, a serotonin 5-HT 1B/1D receptor ligand and drug for migraine treatment [118].Autoradiography on rhesus monkey brain tissue revealed selective binding (90%) to high-affinity binding sites and the tracer was blockable by 5-HT 1B and 5-HT 1D but not by 5-HT 1A receptor antagonist.The compound was also used in human PET studies [119,120], as was the isotopomer [methyl- 11 C]Zolmitriptan, Scheme 19 [121].
Dahl et al expanded the early carbamate work performed by Doi et al by labelling the first thiocarbamate (61%) using a thiol as a nucleophile [28] and Stevens et al synthesized [ 11 C]sulfonyl carbamates by employing sulfonyl azides previously employed in sulfonylurea labelling.Thirteen compounds were synthesized (3-88%) plus a nonpeptide angiotensin II receptor subtype 2 agonist 58 (23%), which was evaluated by a human prostate carcinoma cell assay, ex vivo biodistribution and PET-imaging in rat [128].
Antiepileptic [ 11 C]Phenytoin 40 (22%) carrying a hydantoin structure was synthesized via carbonylative intramolecular ring closure of an azide and amide group with high molar activity, Scheme 20 [27].The product was obtained in higher radiochemical yield compared to the first reported synthesis based on [ 11 C]phosgene [17].The compound underwent preclinical [129] and clinical [130,131] evaluation as a P-gp tracer.In preclinical studies the compound behaved as a moderate Pgp substrate in small animal PET, had high metabolic stability in the brain and the baseline signal in brain was higher compared to other Pgp tracers.

Nickel-mediated 11 C-carbonylation
While Pd-mediated carbonylation gives access to a wide array of 11 Clabelled compounds the scope is restricted to organic halides or triflates that are lacking β-hydrogens, e.g.methyl, aryl, benzyl and alkenyl iodides.β-Hydride elimination is known to be less prone to occur in nickel-relative to palladium-catalysed coupling reactions, attributed to differences in activation energy for this side-reaction [132].Recently, 11 C-carbonylative cross-coupling of β-hydrogen containing cyclohexyl, propyl and pentyl iodides with aminopyridine (43-72%) and benzylamine (33-25%) was accomplished using air sensitive Ni (COD) 2 -complex with bathophenanthroline as supporting ligand.[133] Hence, the synthetic scope for transition-metal mediated 11 C-carbonylation had been considerably widened.First practically useful free radical-mediated carbonylation protocols were developed in the late 1990s as a convenient route to aliphatic carboxylic amides and esters [134,135].Like transition metal-mediated carbonylation it is a three-component coupling reaction, typically of an alkyl iodide, CO and an amine, or another nucleophile.Unlike transition metal-mediated carbonylation this route is essentially free from the β-hydride elimination side-reaction.The mechanism of radicalmediated carbonylation is understood to proceed through several steps leading to the formation of an acyl iodide intermediate.The acyl iodide further reacts with a nucleophile to yield the carbonyl compound.In practice, the radical chain reaction is induced by cleavage of the carbon-iodine bond by UV-irradiation of the reaction mixture or by thermal radical initiators, the latter being the more common method, perhaps for its simplicity.

UV-initiated radical 11 C-carbonylation
The protocol for free radical carbonylation with [ 11 C]CO was developed by Itsenko et al using a high-pressure (35 MPa) autoclave equipped with a sapphire window to enable UV irradiation.Performing the reactions in the high-pressure autoclave was an important part of the protocol since the reagent solutions did not trap any [ 11 C]CO if bubbled through the reaction mixture.Even the most reactive reactants failed to convert [ 11 C]CO into product.The explanation pointed to the reversibility of the addition of CO to an alkyl radical.
The first report included the syntheses of twelve [carbonyl-11 C] carboxamides using predominantly primary and secondary alkyl iodides and amines of varying nucleophilicity [136].Reactions were favoured by polar aprotic solvents such as N-methyl-2-pyrrolidone (NMP) and 1,3-dimethylimidazolidin-2-one (DMI).Radiochemical yields up to 79% were obtained with primary and secondary alkyl iodides and more reactive amines, while tert-butyl iodide provided modest 20%.The commonly used substrates in transition metal-mediated carbonylations, iodomethane gave a similar result while iodobenzene only gave 4% yield.Low yields were also obtained in syntheses with less nucleophilic amines.
Addition of a strong base (KOH, BuLi, TBAOH or LiHDMS) capable of deprotonating the hydroxy group was necessary to obtain useful radiochemical yields of [ 11 C]carboxylic esters (40-67%) [137].Reactions were performed in pure alcohols or in THF-alcohol solvent mixtures.
The radical 11 C-carbonylation method was further extended to [1-11 C]carboxylic acid synthesis [138,139].As with other oxygen nucleophiles the reaction required a base, such as TBAOH and KOH to be added to THF-water or acetonitrile-water based reaction mixtures.Radiochemical yields up to 70% were obtained with primary and secondary alkyl iodides.tert-Butyl iodide could not be carboxylated, which was attributed to the extra stability of tert-butyl radical.Tertiary 1-iodoadamanane, in contrast, provided the corresponding acid in 63% radiochemical yield.Some of the model compounds included functionalised acids that would not be accessible through the direct reaction of Grignard regents with [ 11 C]CO 2 .
The bases used for enhancing the reactivity of oxygen nucleophiles in the synthesis of 11 C-labelled carboxylic acids and esters was successfully replaced by photosensitisers, thus enabling equally good radiochemical yields at base-free conditions [140,141].The milder reaction conditions may have helped minimising competing reactions, e.g.direct nucleophilic substitution between the two reaction substrates.
[1- 11 C]Carboxylic acids could also be obtained in good yields simply by UV-irradiation of alkyl iodides in DMSO at 35 °C [142].Synthesized among other compounds was [ 11 C]Valproic acid 11 (75%), a registered drug for in therapy of epilepsy, bipolar disorders and migraine, Scheme 21.The sulfoxide oxygen was supposed to act as an oxygen nucleophile in anhydrous conditions.

Thermally initiated radical 11 C-carbonylation
A protocol for thermally initiated radical 11 C-aminocarbonylation of alkyl iodides in glass vials was presented by Chow et al utilising [ 11 C]CO transfer in xenon gas [61].AIBN as radical initiator and low-pressure delivery of [ 11 C]CO represents an attractive complement to the previous radical 11 C-carbonylation method that used a UV-light source and a high pressure reactor.Seven labelled amides were obtained with high reproducibility and decay corrected isolated radiochemical yields between 9 and 25%, including the 11β-HSD1 inhibitor [ 11 C]adamantan-1-yl(piperidin-1-yl)methanone 34 (18%), Scheme 22. [carbonyl- 11 C]N-Phenylcyclohexanecarboxamide (20%) was isolated with 1.32 GBq, >99% radiochemical purity and a molar activity of 101 ± 13 GBq/ μmol.In addition, one carboxylic ester (11%) was labelled by replacing the amine nucleophile with isopropanol.

Mechanistic studies relating to [ 11 C]CO
[1- 11 C]Ethyl chloride produced from [ 11 C]CO was used to study kinetic isotope effects in the S N 2 reaction between ethyl chloride and cyanide ions, Scheme 23 [143,144].The labelled ethyl chloride was obtained by 11 C-hydroxycarbonylation of methyl iodide, reduction to [1- 11 C]ethanol and treatment with concentrated hydrochloric acid and anhydrous ZnCl 2 .Mechanistic studies on [ 11 C]methanol transformations by alumina, zeolite catalysts [145], vanadium V-MCM-41 mesoporous silica [146,147] and copper and chromium modified SBA-15 mesoporous silica [148] demonstrated formation of [ 11 C]CO among other compounds.methods that can provide new tracer candidates is one of the compelling aspects of radiochemistry research.The compounds presented in Table 2 are examples of biologically active structures that have been synthesized using 11 C-carbonylation methods and demonstrates the structural diversity and potential that lays in this chemistry.

Biologically evaluated compounds synthesized from [ 11 C]CO
While 11 C-carbonylation chemistry has been practiced by a relatively small number of research groups, and thus not yet been implemented on a wider scale in the PET community, several labelled compounds have been biologically evaluated.Fig. 5 visualizes the number of studies that have been published, although it is probably only a fraction of the actual number, since all work is not public or publishable.The 11 Clabelled compounds used in the studies are presented in Table 3.The label was incorporated at the lactone ring by Pd-mediated reaction using the high-pressure autoclave carbonylation method, Fig. 1.

Clinical applications
However, to the best of our knowledge, the first clinical PET scan with a PET-tracer labelled by 11 C-carbonylation was performed in 2000 with the NMDA glycine site antagonist [ 11 C]GV15026 41.GV15026 was developed as an anti-stroke drug.A PET study in healthy volunteers failed to demonstrate uptake in brain parenchyma, hence the compound was not suitable as an in vivo tracer for the glycine receptor (previously unpublished results).Another example from the PET facility in Uppsala is [carbonyl-11 C]Zolmitriptan 49, labelled in the oxazolidinone ring by selenium-mediated carbonylation.The deposition and distribution of [carbonyl- 11 C]Zolmitriptan and an isotopomer labelled in the N-methyl position [121] were evaluated after intranasal administrations.The relation between brain and plasma drug concentration was assessed after an intravenous PET-tracer administration combined with intranasal Zolmitriptan drug administration to establish a therapeutic relevant plasma drug concentration [119,120].The intranasal route of administration indicated drug uptake through nasal mucosa contributing to rapid systemic availability.From the point of view of production, the 11 C-carbonylation method simplified the process due to easier separation of the product from the precursor compared to the 11 C-methylation method.
In recent publications by Mansor et al, the P-gp tracer [ 11 C]Phenytoin 40 was examined in six healthy volunteers who were scanned two times the same day with arterial blood sampling [130,131].Singletissue-compartment model with blood volume parameter was identified as a good model for quantification of dynamic [ 11 C]Phenytoin PET.The synthesis method was initially developed for preclinical imaging before being validated for GMP compliant manufacturing [129].The validation included ICP analysis of the product solution which showed insignificant concentrations of rhodium (ppb), hence it was concluded that the rhodium was efficiently removed during the purification process.

Good manufacturing practice and PET-tracer production
Good manufacturing practise (GMP) is essential for a safe and reproducible production of PET-tracers.The aim of GMP for radiopharmaceuticals can be formulated as follows: the product is delivered at the right time, in sufficient amount and quality for the intended purpose.The process of implementing this paradigm to the current level has required substantial efforts, not only by radiochemists but also from authorities to create a modified framework of GMP coined "PET-GMP".This simplified regulation takes several restrictions into account, e.g. the short time from production to patient use due to the short physical half-lives of the radionuclides, the very small amount of active substance in the product (few micrograms) and need for radiation safety when handling high levels of radioactivity.The level enforced by PET-GMP today is substantially higher compared to the situation in the 1980s when PET-tracer production could be performed in radiochemistry laboratories that were not built for synthesis of parenteral radiopharmaceuticals.Further improvement in PET-GMP will most likely be implemented in the future and new synthesis technology might be a key factor helping the PETcommunity to adhere to the demands of regulatory authorities.The four 11 C-carbonylation PET-tracers administrated in man, [ 11  Although quality control always precedes release of radiopharmaceutical products, the use of consumables reduces the risks associated with carry-over and simplify the validation of production processes.More suitable techniques for GMP-production than the autoclave method, are low-pressure methods that facilitate use of single-use reaction vessels and straight forward automation, e.g.xenon gas to transfer [ 11 C] CO to the reaction mixture or, if applicable for the synthesis, the use of Pd-Xantphos complexes for efficient [ 11 C]CO capture [60].When producing tracers according to GMP, most aspects surrounding 11 C-carbonylations are identical to those of common 11 C-methylations, e.g. the radionuclide production, the purification of the labelled compounds, the reformulation and the routine quality control which includes sterility, endotoxin content, pH, radiochemical and chemical purity, etc.One special recommendation for Pd and Rh-mediated synthesis is to include trace metal analysis as part of the validation process.However, standard semi-preparative HPLC has shown to efficiently remove both Rh and Pd down to ppb levels in the final product solution, thus no additional purification steps have been necessary.Hence, key factors when establishing GMP production of 11 C-carbonylation-based tracers is the synthesis equipment, together with thorough validation of the synthesis method for maximum reproducibility and delivery reliability.As previously discussed, there are not yet any commercially available synthesis modules for 11 C-carbonylations and GMP production.This could naturally be a reason that so few PET-tracers synthesized from [ 11 C]CO have been administered in human, especially considering the otherwise very versatile chemistry.If this obstacle can be overcome, either through the launch of commercial 11 C-carbonylation equipment or simpler protocols with existing 11 C-synthesis equipment, the next important step would be a successful tracer that gives strong incentives to implement capability for [ 11 C]CO 2 , [ 11 C]CO, [ 11 C]methyl iodide, [ 11 C] methyl triflate and ammonium [ 11 C]cyanide [ 11 C]CO from [ 11 C]CO 2 , all

Fig. 1 .
Fig.1.Schematic drawing of a high-pressure system for 11 C-carbonylation equipped with one additional injection valve for introduction of reagent gas, e.g.hydrogen gas for hydroformylation reactions or ( 13 C)CO for co-labelling experiments[45].

11 C-Fig. 2 .
Fig.2.Schematic drawing of a system utilising xenon transfer gas for efficient use of [ 11 C]CO in carbonylation reactions[61].

Fig. 3 .Fig. 4 .
Fig. 3. Number of published compounds synthesized by 11 C-carbonylation, arranged according to their class and principal reaction mediator.

11 C
-carbonylation due to the large excess of both the boronic acid and the active Pd(0)-complex in relation to [ 11 C]CO.[ 11 C]Benzophenone (37%) was recently synthesized with similar reaction conditions[28].
So far there are only a few examples of clinical PET-studies with tracers labelled by 11 C-carbonylation.The first paper was published in 2006 by Erondu et al and described a receptor occupancy study with [ 11 C]MK-0233 43, a selective neuropeptide Y Y5 receptor ligand [108].
C]CO-chemistry.Making this a reality is arguably one of the most important challenges for the research on 11 C-carbonylation chemistry.
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